Real-Time In Vivo Assessment of the Nerve Microenvironment with Coherent Anti–Stokes Raman Scattering Microscopy

Henry, Francis P.; Côté, Daniel; Randolph, Mark A.; Rust, Esther A. Z.; Redmond, Robert W.; Kochevar, Irene E.; Lin, Charles P.; Winograd, Jonathan M.

doi:10.1097/prs.0b013e318191c5b8

Cited by 39 publications

(38 citation statements)

References 10 publications

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“…Nonetheless, CARS imaging has been used to visualize normal myelin and demyelination in a chemical model of demyelination 6,7 and Wallerian degeneration following crush injuries. 25 In MS and EAE, demyelination and axonal dysfunction are associated with inflammatory lesions. 11 Thus, it would be advantageous to image alterations simultaneously in axons, myelin, and immune cells.…”

Section: Discussionmentioning

confidence: 99%

Multimodal coherent anti-Stokes Raman scattering microscopy reveals microglia-associated myelin and axonal dysfunction in multiple sclerosis-like lesions in mice

et al. 2011

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Abstract. Myelin loss and axonal degeneration predominate in many neurological disorders; however, methods to visualize them simultaneously in live tissue are unavailable. We describe a new imaging strategy combining video rate reflectance and fluorescence confocal imaging with coherent anti-Stokes Raman scattering (CARS) microscopy tuned to CH 2 vibration of myelin lipids, applied in live tissue of animals with chronic experimental autoimmune encephalomyelitis (EAE). Our method allows monitoring over time of demyelination and neurodegeneration in brain slices with high spatial resolution and signal-to-noise ratio. Local areas of severe loss of lipid signal indicative of demyelination and loss of the reflectance signal from axons were seen in the corpus callosum and spinal cord of EAE animals. Even in myelinated areas of EAE mice, the intensity of myelin lipid signals is significantly reduced. Using heterozygous knock-in mice in which green fluorescent protein replaces the CX 3 CR1 coding sequence that labels central nervous system microglia, we find areas of activated microglia colocalized with areas of altered reflectance and CARS signals reflecting axonal injury and demyelination. Our data demonstrate the use of multimodal CARS microscopy for characterization of demyelinating and neurodegenerative pathology in a mouse model of multiple sclerosis, and further confirm the critical role of microglia in chronic inflammatory neurodegeneration. C 2011 Society of Photo-Optical Instrumentation Engineers (SPIE).

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Section: Discussionmentioning

confidence: 99%

Multimodal coherent anti-Stokes Raman scattering microscopy reveals microglia-associated myelin and axonal dysfunction in multiple sclerosis-like lesions in mice

et al. 2011

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“…Second, CARS imaging does not require labeling for visualization; thus, it allows long-term visualization of the myelin sheaths ( 41,115 ). Third, CARS-based multimodal microscopy allows simultaneous visualization of myelin sheaths and cellular processes during demyelination (111)(112)(113). This capability is invaluable to the studies of molecular mechanisms underlying myelin diseases.…”

Section: Study Of Lipids In Cancer Developmentmentioning

confidence: 99%

“…This capability is invaluable to the studies of molecular mechanisms underlying myelin diseases. Fourth, CARS microscopy allows in vivo imaging of the myelin sheaths ( 43,44,113 ). This capability is invaluable for the investigation of the causes of myelin diseases or the evaluation of drugs aiming at curtailing further demyelination or restoring the structure and function of the myelin sheaths.…”

Section: Study Of Lipids In Cancer Developmentmentioning

confidence: 99%

Shedding new light on lipid biology with coherent anti-Stokes Raman scattering microscopy

Yue

Cheng

2010

Journal of Lipid Research

147

130

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Despite the ubiquitous roles of lipids in biology, the detection of lipids has relied on invasive techniques and population measurements. The molecular profi les of the lipid-rich structures are evaluated by measurements of total lipid extracts with liquid or gas chromatography coupled to mass spectrometry ( 8 ). This approach is inexact because it measures lipid molecules from areas other than the structures of interest. Furthermore, spatial information, which is critical to understanding the function of lipids ( 9 ), is inaccessible with the bulk measurement techniques. To visualize the lipid-rich structures, fl uorescently tagged lipid molecules, which intercalate with the lipidrich structures, are used ( 10, 11 ). Nevertheless, the use of the fl uorescent lipid molecules to label lipid-rich structures is problematic for several reasons. First, to facilitate the transport of the fl uorescent lipid molecules into cells, cell fi xation procedures are often performed ( 12 ). This procedure prevents monitoring the dynamic responses of lipid-rich structures to stimuli or the disease processes. Second, labeling effi ciency is highly dependent on the type of fl uorescent lipid molecules ( 12 ), thus posing a signifi cant problem to the quantitation of lipid-rich structures. Third, the intercalation of fl uorescent lipid molecules can lead to changes in the properties of lipidrich structures. For instance, incorporation of fl uorescent lipid molecules into the cell membrane can induce membrane phase separation and membrane microdomain formation, which can perturb critical cellular processes including signal transduction and endocytosis ( 13 ).As an alternative to fl uorescence, signals from molecular vibration provide an attractive means for label-free chemical imaging. In particular, Raman scattering has been widely used for spectroscopic study of biomolecules ( 14 ). The Raman-scattered photons exhibit frequency Abstract Despite the ubiquitous roles of lipids in biology, the detection of lipids has relied on invasive techniques, population measurements, or nonspecifi c labeling. Such diffi culties can be circumvented by a label-free imaging technique known as coherent anti-Stokes Raman (CARS) microscopy, which is capable of chemically selective, highly sensitive, and high-speed imaging of lipid-rich structures with submicron three-dimensional spatial resolution. We review the broad applications of CARS microscopy to studies of lipid biology in cell cultures, tissue biopsies, and model organisms. Recent technical advances, limitations of the technique, and perspectives are discussed. Lipids play a critical role in human health and diseases. Phospholipids, glycolipids, and sterol lipids are the major components of the cell membrane ( 1 ). Sphingolipids constitute up to 80% of the myelin sheaths, which are the membranous structures that wrap around the axons for insulation and for proper propagation of electrical impulses ( 2 ). Glycerolipids or triglycerides serve as the cytoplasmic energy depots ( 3 ). Bioactive lip...

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“…Both CARS and SHG are emitted in the epidirection if the particles of interest are subwavelength; however, the forward propagating signal also rescatters in the backward direction and represents the majority of the signal collected [44,51]. Given the large lipid content in myelin, CARS has been particularly successful in studying nerve damage and recovery [52][53][54]. When imaging in live animals, longitudinal studies are possible to better understand damage to the spine, coping mechanisms, and long-term recovery.…”

Section: Applicationsmentioning

confidence: 99%

Single laser source for multimodal coherent anti-Stokes Raman scattering microscopy

et al. 2010

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Real-Time In Vivo Assessment of the Nerve Microenvironment with Coherent Anti–Stokes Raman Scattering Microscopy

Cited by 39 publications

References 10 publications

Multimodal coherent anti-Stokes Raman scattering microscopy reveals microglia-associated myelin and axonal dysfunction in multiple sclerosis-like lesions in mice

Multimodal coherent anti-Stokes Raman scattering microscopy reveals microglia-associated myelin and axonal dysfunction in multiple sclerosis-like lesions in mice

Shedding new light on lipid biology with coherent anti-Stokes Raman scattering microscopy

Single laser source for multimodal coherent anti-Stokes Raman scattering microscopy

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